Tissue factor pathway inhibitor (TFPI) is 276 amino acids in length and functions as an inhibitor of tissue factor-mediated blood coagulation. Its amino acid sequence is shown in SEQ ID NO:1. The amino terminal end of TFPI is negatively charged, and the carboxy terminal end is positively charged. The TFPI protein contains three Kunitz-type enzyme inhibitor domains. TFPI contains 18 cysteine residues and forms 9 disulfide bridges when correctly folded. The primary sequence contains three N-linked consensus glycosylation sites (Asn-X-Ser/Thr). The asparagine residues of the glycosylation sites are located at positions 145, 195, and 256. TFPI is also known as lipoprotein associated coagulation inhibitor (LACI), tissue factor inhibitor (TFI), and extrinsic pathway inhibitor (EPI).
Use of TFPI has been proposed for the treatment of various indications, including sepsis (U.S. Pat. No. 6,063,764 and WO 93/24143), deep vein thrombosis (U.S. Pat. No. 5,563,123, U.S. Pat. No. 5,589,359, and WO 96/04378), ischemia (U.S. Pat. No. 5,885,781, U.S. Pat. No. 6,242,414, and WO 96/40224), restenosis (U.S. Pat. No. 5,824,644 and WO 96/01649), and cancer (U.S. Pat. No. 5,902,582 and WO 97/09063). A TFPI variant, which differs from TFPI by the addition of an alanine residue at the amino terminus (“ala-TFPI”), has been shown to be efficacious in animal models for the treatment of sepsis. Carr et al., Circ Shock November 1994; 44(3):126-37.
Following preparation, the compositions of TFPI or a TFPI variant can be packaged for storage in an aqueous form or in a frozen state. TFPI or TFPI variants, however, can form aggregates during storage in aqueous formulations. Aggregation is caused by interactions between the TFPI or TFPI variant molecules that result in the formation of oligomers. These oligomers may remain soluble or may form large, visible aggregates that precipitate from solution during storage. Aggregate formation by TFPI or TFPI variant during storage of an aqueous composition can adversely affect its biological activity, resulting in loss of therapeutic efficacy as an anti-coagulant effective for the treatment of a variety of conditions, including sepsis. Furthermore, aggregate formation may cause other problems, such as blockage of tubing, membranes, or pumps when the TFPI-containing or TFPI variant-containing composition is administered using an infusion system. To minimize these problems, there is a need in the art for improved stabilization of compositions of TFPI and TFPI variants.